The leukotrienes constitute a relatively new group of biologically active compounds initially derived from arachidonic acid oxygenated at C-5 in a lipoxygenase catalyzed reaction. The study of these compounds has undergone explosive growth since the description of LTC4 as a slow reacting substance of anaphylaxis due to potential for rapid application of new basic knowledge to the clinical situation. The analytical methodology available to quantitate leukotrienes is somewhat limited, in particular the availability of reference physical chemical technique to validate bioassay or the RIA technique. In this application, we propose to develop sophisticated mass spectrometric techniques for sensitive, precise, and accurate quantitation of 5-HETE, LTB4, LTC4, LTD4 and LTE4 following fundamental studies of negative ion chemical ionization mass spectrometry of pentafluorobenzyl esters of the hydroxy fatty acid lipoxygenase metabolites. Desulfurization and hydrogenation of the sulfido-leukotrienes to 5-hydroxyeicosanoic acid is proposed as a means to sensitively quantitate the myotropic LTs by mass spectrometry. Multiple UV scanning techniques will also be evaluated as a means to obtain characteristic UV spectra from a few nanograms of LTC4. Physicochemical methods developed in this program will be applied to the study of leukotriene biosynthesis in isolated cells stimulated by hypoxia and in clinical samples of lung lavagates from patients with pulmonary hypertension due to various pathophysiological causes. Additional studies are proposed to investigate the chemical degradation of LTC4/D4/E4 catalyzed by iron salts as well as the degradation of LTB4 by hydroxyl and hydroperoxide radical. Investigation of the in vitro and in vivo metabolism of leukotrienes as well as 5-HETE is proposed with structure elucidation of major metabolites as a prime goal. Finally, the chemical structure of an antimyotropic substance (AMS) isolated from lung lavagates which we have recently discovered, will be sought. This substance has been found in the lung of rats treated with monocrotaline in a model of chronic pulmonary hypertension. Human AMS will also be studied in the lavagates of patients with persistent pulmonary hypertension of the newborn. This antimyotropic substance appears to be synthesized in response to chronic pulmonary vasoconstriction and may be an important, previously unrecognized, mediator in pulmonary vascular homeostasis.